title: Step-by-step mechanism of DNA damage recognition by human 8-oxoguanine DNA glycosylase.
abstract: BACKGROUND
  Extensive structural studies of human DNA glycosylase hOGG1 have revealed essential conformational changes of the enzyme. However, at present there is little information about the time scale of the rearrangements of the protein structure as well as the dynamic behavior of individual amino acids.
  
  
  METHODS
  Using pre-steady-state kinetic analysis with Trp and 2-aminopurine fluorescence detection the conformational dynamics of hOGG1 wild-type (WT) and mutants Y203W, Y203A, H270W, F45W, F319W and K249Q as well as DNA-substrates was examined.
  
  
  RESULTS
  The roles of catalytically important amino acids F45, Y203, K249, H270, and F319 in the hOGG1 enzymatic pathway and their involvement in the step-by-step mechanism of oxidative DNA lesion recognition and catalysis were elucidated.
  
  
  CONCLUSIONS
  The results show that Tyr-203 participates in the initial steps of the lesion site recognition. The interaction of the His-270 residue with the oxoG base plays a key role in the insertion of the damaged base into the active site. Lys-249 participates not only in the catalytic stages but also in the processes of local duplex distortion and flipping out of the oxoG residue. Non-damaged DNA does not form a stable complex with hOGG1, although a complex with a flipped out guanine base can be formed transiently.
  
  
  CONCLUSIONS
  The kinetic data obtained in this study significantly improves our understanding of the molecular mechanism of lesion recognition by hOGG1.
authors.affiliation: Institute of Chemical Biology and Fundamental Medicine, Siberian Branch of the Russian Academy of Sciences, Novosibirsk 630090, Russia; Department of Natural Sciences, Novosibirsk State University, Novosibirsk 630090, Russia.
authors.forename: Alexandra A
authors.initials: AA
authors.lastname: Kuznetsova
authors.forename: Nikita A
authors.initials: NA
authors.lastname: Kuznetsov
authors.forename: Alexander A
authors.initials: AA
authors.lastname: Ishchenko
authors.forename: Murat K
authors.initials: MK
authors.lastname: Saparbaev
authors.forename: Olga S
authors.initials: OS
authors.lastname: Fedorova
mesh_terms: D001665:Binding Sites; D002384:Catalysis; D004249:DNA Damage / Q000235:genetics*; D045647:DNA Glycosylases / Q000235:genetics / Q000378:metabolism*; D004260:DNA Repair; D006147:Guanine / Q000031:analogs & derivatives* / Q000378:metabolism; D006801:Humans; D007700:Kinetics; D008968:Molecular Conformation; D050505:Mutant Proteins / Q000235:genetics / Q000378:metabolism*; D009154:Mutation / Q000235:genetics; D013050:Spectrometry, Fluorescence
chemical_list: D050505:Mutant Proteins; C024829:8-hydroxyguanine; D006147:Guanine; D045647:DNA Glycosylases; C479058:oxoguanine glycosylase 1, human
keywords: (3-hydroxytetrahydrofuran-2-yl)methyl phosphate; 2-aPu; 2-aminopurine; 7,8-dihydro-8-oxoguanine; BER; Base excision repair; Conformational dynamics; DNA glycosylase; Enzyme kinetics; Human 8-oxoguanine DNA glycosylase; ODN; THF; base excision repair; oligodeoxyribonucleotide; oxoG